ANALYSIS OF PHYTATE CONTENT VARIABILITY AND MOLECULAR CHARACTERISATION OF GENES IN COWPEA LANDRACES

Abstract

Phytate is a naturally occurring phosphorus-storage compound in seeds, but it reduces the bioavailability of iron, zinc, and calcium in human diets. Cowpea (Vigna unguiculata (L.) Walp), an essential dietary protein source widely consumed in Nigeria, contains notable phytate levels, which may hinder micronutrient absorption with regular consumption. This study investigated phytate content in Nigerian cowpea accessions and explored candidate genes involved in its biosynthesis to guide future efforts for nutritional improvement. Thirty (30) cowpea landraces were obtained from the National Centre for Genetic Resources and Biotechnology (NACGRAB). Phytate concentration was quantified using acid digestion followed by filtration and titration. To explore the genetic basis of phytate accumulation, protein sequences of known phytate biosynthetic genes from Arabidopsis thaliana, Glycine max, and Phaseolus vulgaris were retrieved from Phytozome 14.0. These sequences were used for BLASTp searches to identify putative homologues in cowpea. Homologous cowpea sequences were aligned and subjected to phylogenetic analysis using the Neighbour-Joining method in MEGA v12.0.11 with 10,000 bootstrap replicates to assess evolutionary relationships. The results showed significant variation in phytate content, 3.96‒30.94mg/g (p < 0.001) across cowpea accessions. The cowpea sequences displayed strong homology to known phytate biosynthetic enzymes in seed, MIPS (Myo-inositol-3-phosphate synthase), ITPK (Inositol 1,3,4-triphosphate 5/6-kinase), IPK2 (Inositol 1,4,5-tris-phosphate kinase), MRP (Multidrug-resistance-associated protein ATP-binding cassette), IMP (Inositol monophosphate phosphatase), MIK (Myo-inositol kinase), and IPK1 (Inositol 1,3,4,5,6 pentakisphosphate 2-kinase), with low E-values and high identity percentages. Phylogenetic clustering of cowpea sequences alongside those from related legumes suggests potential functional conservation, although expression-level validation is necessary. Quantitative gene expression profiling to further clarify the roles of these genes in phytate biosynthesis and accumulation is needed. These identified genes could be prioritised for reverse genetics or transcriptomic studies, provided that tissue-specific expression and off-target impacts are carefully evaluated